SARS-CoV-2 NSP7 inhibits type I and III IFN production by targeting the RIG-I/MDA5, TRIF, and STING signaling pathways.

SARS-CoV-2 is a poor inducer of innate antiviral immunity, and the underlying mechanism still needs further investigation. Here, we reported that SARS-CoV-2 NSP7 inhibited the production of type I and III IFNs by targeting the RIG-I/MDA5, TLR3-TRIF, and cGAS-STING signaling pathways. SARS-CoV-2 NSP7 suppressed the expression of IFNs and IFN-stimulated genes induced by poly (I:C) transfection and infection with Sendai virus or SARS-CoV-2 virus-like particles. NSP7 impaired type I and III IFN production activated by components of the cytosolic dsRNA-sensing pathway, including RIG-I, MDA5, and MAVS, but not TBK1, IKKε, and IRF3-5D, an active form of IRF3. In addition, NSP7 also suppressed TRIF- and STING-induced IFN responses. Mechanistically, NSP7 associated with RIG-I and MDA5 prevented the formation of the RIG-I/MDA5-MAVS signalosome and interacted with TRIF and STING to inhibit TRIF-TBK1 and STING-TBK1 complex formation, thus reducing the subsequent IRF3 phosphorylation and nuclear translocation that are essential for IFN induction. In addition, ectopic expression of NSP7 impeded innate immune activation and facilitated virus replication. Taken together, SARS-CoV-2 NSP7 dampens type I and III IFN responses via disruption of the signal transduction of the RIG-I/MDA5-MAVS, TLR3-TRIF, and cGAS-STING signaling pathways, thus providing novel insights into the interactions between SARS-CoV-2 and innate antiviral immunity. This article is protected by copyright. All rights reserved.

Doctors' and Patients' Perceptions of Impacts of Doctors' Communication and Empathy Skills on Doctor-Patient Relationships During COVID-19.

BACKGROUND: During the COVID-19 pandemic, the performance of Chinese doctors may have led to improved doctor-patient relationships (DPRs). However, it is unclear how doctors and patients perceived the impact of doctors' communication and empathy skills on DPRs during the COVID-19 pandemic. OBJECTIVE: To examine the perceptions of doctors and patients on how doctors' communication skills and empathy skills influence DPRs during COVID-19. MAIN MEASURES: Doctors' and patients' perceptions of doctors' communication skills were measured using the Chinese version of the SEGUE Framework. To measure empathy skills and DPRs, the Jefferson Scale of Empathy and Difficult Doctor-Patient Relationship Questionnaire were administered to doctors, and the Consultation and Relational Empathy Measure and Patient-Doctor Relationship Questionnaire were administered to patients. RESULTS: A total of 902 doctors and 1432 patients in China were recruited during the pandemic via online or offline surveys (overall response rate of 69.8%). Both doctors and patients rated doctors' empathy skills as more impactful on DPRs than communication skills. Doctors believed that only their empathy skills influenced DPRs. But patients believed that there was a significant bi-directional relationship between doctors' communication and empathy skills and these two skills interacted to directly and indirectly influence DPRs, and doctors' empathy had a greater mediating effect than their communication. CONCLUSIONS: During COVID-19, there were both similarities and differences between Chinese doctors' and patients' views on how doctors' communication and empathy skills influenced DPRs. The greater effect of doctors' empathy skills suggests that both doctors and patients attach more importance to doctors' empathy in doctor-patient interactions. The bi-directional effect on patient outcomes suggests that both doctors' communication and empathy skills are important to patients' perceptions of DPRs.

Manipulation of innate immune signaling pathways by SARS-CoV-2 non-structural proteins.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of the current coronavirus disease 2019 (COVID-19) pandemic, induces an unbalanced immune response in the host. For instance, the production of type I interferon (IFN) and the response to it, which act as a front-line defense against virus invasion, are inhibited during SARS-CoV-2 infection. In addition, tumor necrosis factor alpha (TNF-α), a proinflammatory cytokine, is upregulated in COVID-19 patients with severe symptoms. Studies on the closely related betacoronavirus, SARS-CoV, showed that viral proteins such as Nsp1, Orf6 and nucleocapsid protein inhibit IFN-ß production and responses at multiple steps. Given the conservation of these proteins between SARS-CoV and SARS-CoV-2, it is not surprising that SARS-CoV-2 deploys similar immune evasion strategies. Here, we carried out a screen to examine the role of individual SARS-CoV-2 proteins in regulating innate immune signaling, such as the activation of transcription factors IRF3 and NF-κB and the response to type I and type II IFN. In addition to established roles of SARS-CoV-2 proteins, we report that SARS-CoV-2 proteins Nsp6 and Orf8 inhibit the type I IFN response but at different stages. Orf6 blocks the translocation of STAT1 and STAT2 into the nucleus, whereas ORF8 inhibits the pathway in the nucleus after STAT1/2 translocation. SARS-CoV-2 Orf6 also suppresses IRF3 activation and TNF-α-induced NF-κB activation.

Evolutionary plasticity of zoonotic porcine Deltacoronavirus (PDCoV): genetic characteristics and geographic distribution.

The emergence and rapid spread of the acute respiratory syndrome coronavirus-2 have confirmed that animal coronaviruses represent a potential zoonotic source. Porcine deltacoronavirus is a worldwide evolving enteropathogen of swine, detected first in Hong Kong, China, before its global identification. Following the recent detection of PDCoV in humans, we attempted in this report to re-examine the status of PDCoV phylogenetic classification and evolutionary characteristics. A dataset of 166 complete PDCoV genomes was analyzed using the Maximum Likelihood method in IQ-TREE with the best-fitting model GTR + F + I + G4, revealing two major genogroups (GI and GII), with further seven and two sub-genogroups, (GI a-g) and (GII a-b), respectively. PDCoV strains collected in China exhibited the broadest genetic diversity, distributed in all subgenotypes. Thirty-one potential natural recombination events were identified, 19 of which occurred between China strains, and seven involved at least one China strain as a parental sequence. Importantly, we identified a human Haiti PDCoV strain as recombinant, alarming a possible future spillover that could become a critical threat to human health. The similarity and recombination analysis showed that PDCoV spike ORF is highly variable compared to ORFs encoding other structural proteins. Prediction of linear B cell epitopes of the spike glycoprotein and the 3D structural mapping of amino acid variations of two representative strains of GI and GII showed that the receptor-binding domain (RBD) of spike glycoprotein underwent a significant antigenic drift, suggesting its contribution in the genetic diversity and the wider spread of PDCoV.

Rapid and Sensitive Genotyping of SARS-CoV-2 Key Mutation L452R with an RPA-PfAgo Method.

In the two years of COVID-19 pandemic, the SARS-CoV-2 variants have caused waves of infections one after another, and the pandemic is not ending. The key mutations on the S protein enable the variants with enhanced viral infectivity, immune evasion, and/or antibody neutralization resistance, bringing difficulties to epidemic prevention and control. In support of precise epidemic control and precision medicine of the virus, a fast and simple genotyping method for the key mutations of SARS-CoV-2 variants needs to be developed. By utilizing the specific recognition and cleavage property of the nuclease Argonaute from Pyrococcus furiosus (PfAgo), we developed a recombinase polymerase amplification (RPA) and PfAgo combined method for a rapid and sensitive genotyping of SARS-CoV-2 key mutation L452R. With a delicate design of the strategy, careful screening of the RPA primers and PfAgo gDNA, and optimization of the reaction, the method achieves a high sensitivity of a single copy per reaction, which is validated with the pseudovirus. This is the highest sensitivity that can be achieved theoretically and the highest sensitivity as compared to the available SARS-CoV-2 genotyping assays. Using RPA, the procedure of the method is finished within 1.5 h and only needs a minimum laboratorial support, suggesting that the method can be easily applied locally or on-site. The RPA-PfAgo method established in this study provides a strong support to the precise epidemic control and precision medicine of SARS-CoV-2 variants and can be readily developed for the simultaneous genotyping of multiple SARS-CoV-2 mutations.

A DNA-Cu nanocluster and exonuclease I integrated label-free reporting system for CRISPR/Cas12a-based SARS-CoV-2 detection with minimized background signals.

CRISPR-driven biosensing is developing rapidly, but current studies mostly adopt dye-labeled ssDNA as the signal reporter, which is costly and unstable. Herein, we developed a label-free and low-background reporter for CRISPR/Cas12a signaling by integrating DNA-templated copper nanoclusters (DNA-CuNCs) and exonuclease I (EXO I). The template of the DNA-CuNCs was rationally designed as a ds-/ss-DNA hybrid, ensuring that after a quick and nonpersistent cut of Cas12a, a majority of the template can be digested by EXO I. Based on this novel reporter, a biosensor termed CRISPR-CNS (cost-effective, nimble, and sensitive copper nanocluster sensor integrating CRISPR) was developed. Due to the high signal-to-background ratio of our proposed reporter, CRISPR-CNS shows excellent performances for nucleic acid detection, yielding a detection limit of 20 copies for SARS-CoV-2 RNA. Considering its facile synthesis, robust fluorescence, effective cost, and good sensitivity, this combination shall serve as a highly potential output for CRISPR-based point-of-care testing.

Recurrent presence of SARS-CoV-2 RNA in a 33-year-old man

A 33- year-old man had four times recurrent presence of SARS-CoV-2 RNA after 137 days onset of illness, the presence of antibodies did not prevent recurrent infections, but maybe inhibited transmission. This case could have implications for the properties of future vaccines. Reasons for the repeated presence of SARS-CoV-2 RNA may be contribute to the biological characteristics of SARS-CoV-2 and might also be related to coexisting diseases, low neutralizing antibody titres or resistance to antiviral drugs. This article is protected by copyright. All rights reserved.

A Ligation/Recombinase Polymerase Amplification Assay for Rapid Detection of SARS-CoV-2.

The pandemic of COVID-19 caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has led to more than 117 million reported cases and 2.6 million deaths. Accurate diagnosis technologies are vital for controlling this pandemic. Reverse transcription (RT)-based nucleic acid detection assays have been developed, but the strict sample processing requirement of RT has posed obstacles on wider applications. This study established a ligation and recombinase polymerase amplification (L/RPA) combined assay for rapid detection of SARS-CoV-2 on genes N and ORF1ab targeting the specific biomarkers recommended by the China CDC. Ligase-based strategies usually have a low-efficiency problem on RNA templates. This study has addressed this problem by using a high concentration of the T4 DNA ligase and exploiting the high sensitivity of RPA. Through selection of the ligation probes and optimization of the RPA primers, the assay achieved a satisfactory sensitivity of 101 viral RNA copies per reaction, which was comparable to RT-quantitative polymerase chain reaction (RT-qPCR) and other nucleic acid detection assays for SARS-CoV-2. The assay could be finished in less than 30 min with a simple procedure, in which the requirement for sophisticated thermocycling equipment had been avoided. In addition, it avoided the RT procedure and could potentially ease the requirement for sample processing. Once validated with clinical samples, the L/RPA assay would increase the practical testing availability of SARS-CoV-2. Moreover, the principle of L/RPA has an application potential to the identification of concerned mutations of the virus.

Sarcopenia: An underlying treatment target during the COVID-19 pandemic.

The role of skeletal muscle mass in modulating immune response and supporting metabolic stress has been increasingly confirmed. Patients with sarcopenia, characterized by reduced muscle mass and muscle strength, were reported to have poor immune response and metabolic stress when facing acute infection, major surgeries, and other attacks. Based on empirical data, patients with sarcopenia are speculated to have increased infection rates and dismal prognoses amid the current 2019 novel coronavirus disease (COVID-19) epidemic. COVID-19 infection also aggravates sarcopenia because of the increased muscle wasting caused by systematic inflammation and the reduced physical activity and inadequate nutrient intake caused by social isolation. Notably, the interventions targeting skeletal muscle are anticipated to break the vicious circle and benefit the treatment of both conditions. We recommend sarcopenia assessment for populations with advanced age, inactivity, chronic disease, cancers, and nutritional deficiency. Patients with sarcopenia and COVID-19 infection need intensive care and aggressive treatments. The provision of at-home physical activities together with protein supplementation is anticipated to reverse sarcopenia and promote the prevention and treatment of COVID-19. The recommended protocols on nutritional support and physical activities are provided in detail.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) membrane (M) protein inhibits type I and III interferon production by targeting RIG-I/MDA-5 signaling.

Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has quickly spread worldwide and has affected more than 10 million individuals. A typical feature of COVID-19 is the suppression of type I and III interferon (IFN)-mediated antiviral immunity. However, the molecular mechanism by which SARS-CoV-2 evades antiviral immunity remains elusive. Here, we reported that the SARS-CoV-2 membrane (M) protein inhibits the production of type I and III IFNs induced by the cytosolic dsRNA-sensing pathway mediated by RIG-I/MDA-5-MAVS signaling. In addition, the SARS-CoV-2 M protein suppresses type I and III IFN induction stimulated by SeV infection or poly (I:C) transfection. Mechanistically, the SARS-CoV-2 M protein interacts with RIG-I, MAVS, and TBK1, thus preventing the formation of the multiprotein complex containing RIG-I, MAVS, TRAF3, and TBK1 and subsequently impeding the phosphorylation, nuclear translocation, and activation of IRF3. Consequently, ectopic expression of the SARS-CoV-2 M protein facilitates the replication of vesicular stomatitis virus. Taken together, these results indicate that the SARS-CoV-2 M protein antagonizes type I and III IFN production by targeting RIG-I/MDA-5 signaling, which subsequently attenuates antiviral immunity and enhances viral replication. This study provides insight into the interpretation of SARS-CoV-2-induced antiviral immune suppression and illuminates the pathogenic mechanism of COVID-19.

Gastrointestinal involvement attenuates COVID-19 severity and mortality.

Given that gastrointestinal (GI) symptoms are a prominent extrapulmonary manifestation of coronavirus disease 2019 (COVID-19), we investigated intestinal infection with severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) and its effect on disease pathogenesis. SARS-CoV-2 was detected in small intestinal enterocytes by immunofluorescence staining or electron microscopy, in 13 of 15 patients studied. High dimensional analyses of GI tissues revealed low levels of inflammation in general, including active downregulation of key inflammatory genes such as IFNG, CXCL8, CXCL2 and IL1B and reduced frequencies of proinflammatory dendritic cell subsets. To evaluate the clinical significance of these findings, examination of two large, independent cohorts of hospitalized patients in the United States and Europe revealed a significant reduction in disease severity and mortality that was independent of gender, age, and examined co-morbid illnesses. The observed mortality reduction in COVID-19 patients with GI symptoms was associated with reduced levels of key inflammatory proteins including IL-6, CXCL8, IL-17A and CCL28 in circulation but was not associated with significant differences in nasopharyngeal viral loads. These data draw attention to organ-level heterogeneity in disease pathogenesis and highlight the role of the GI tract in attenuating SARS-CoV-2-associated inflammation with related mortality benefit. ONE SENTENCE SUMMARY: Intestinal infection with SARS-CoV-2 is associated with a mild inflammatory response and improved clinical outcomes.

Clinical Features of Fatalities in Patients With COVID-19.

OBJECTIVES: The novel coronavirus disease 2019 (COVID-19) pandemic has spread to over 213 countries and territories. We sought to describe the clinical features of fatalities in patients with severe COVID-19. METHODS: We conducted an Internet-based retrospective cohort study through retrieving the clinical information of 100 COVID-19 deaths from nonduplicating incidental reports in Chinese provincial and other governmental websites between January 23 and March 10, 2020. RESULTS: Approximately 6 of 10 COVID-19 deaths were males (64.0%). The average age was 70.7 ± 13.5 y, and 84% of patients were elderly (over age 60 y). The mean duration from admission to diagnosis was 2.2 ± 3.8 d (median: 1 d). The mean duration from diagnosis to death was 9.9 ± 7.0 d (median: 9 d). Approximately 3 of 4 cases (76.0%) were complicated by 1 or more chronic diseases, including hypertension (41.0%), diabetes (29.0%) and coronary heart disease (27.0%), respiratory disorders (23.0%), and cerebrovascular disease (12.0%). Fever (46.0%), cough (33.0%), and shortness of breath (9.0%) were the most common first symptoms. Multiple organ failure (67.9%), circulatory failure (20.2%), and respiratory failure (11.9%) are the top 3 direct causes of death. CONCLUSIONS: COVID-19 deaths are mainly elderly and patients with chronic diseases especially cardiovascular disorders and diabetes. Multiple organ failure is the most common direct cause of death.

Experts consensus on epidemic prevention and control in radiotherapy centers during the COVID-19 outbreak: Experiences from Sichuan Province.

After the outbreak of COVID-19, medical institutions in China and even around the world are facing unprecedented challenges. In order to minimize the adverse impact of this unexpected epidemic on patients who need radiotherapy, the expert group of our radiotherapy center immediately formulated comprehensive emergency plans and prevention and control measures, partitioned the work area, launched online staff training, and optimized the radiotherapy process after the outbreak, which provided a strong guarantee for the safe and orderly operation of our radiotherapy center and kept the infection rate to an extremely low level. We hope our experience could provide reference and suggestions for other medical institutions.

Increasing host cellular receptor-angiotensin-converting enzyme 2 expression by coronavirus may facilitate 2019-nCoV (or SARS-CoV-2) infection.

The ongoing outbreak of a new coronavirus (2019-nCoV, or severe acute respiratory syndrome coronavirus 2 [SARS-CoV-2]) has caused an epidemic of the acute respiratory syndrome known as coronavirus disease (COVID-19) in humans. SARS-CoV-2 rapidly spread to multiple regions of China and multiple other countries, posing a serious threat to public health. The spike (S) proteins of SARS-CoV-1 and SARS-CoV-2 may use the same host cellular receptor, angiotensin-converting enzyme 2 (ACE2), for entering host cells. The affinity between ACE2 and the SARS-CoV-2 S protein is much higher than that of ACE2 binding to the SARS-CoV S protein, explaining why SARS-CoV-2 seems to be more readily transmitted from human to human. Here, we report that ACE2 can be significantly upregulated after infection of various viruses, including SARS-CoV-1 and SARS-CoV-2, or by the stimulation with inflammatory cytokines such as interferons. We propose that SARS-CoV-2 may positively induce its cellular entry receptor, ACE2, to accelerate its replication and spread; high inflammatory cytokine levels increase ACE2 expression and act as high-risk factors for developing COVID-19, and the infection of other viruses may increase the risk of SARS-CoV-2 infection. Therefore, drugs targeting ACE2 may be developed for the future emerging infectious diseases caused by this cluster of coronaviruses.

Prevalence of depression and its association with quality of life in clinically stable patients with COVID-19.

INTRODUCTION: High risk of mental health problems is associated with Coronavirus Disease 2019 (COVID-19). This study explored the prevalence of depressive symptoms (depression hereafter) and its relationship with quality of life (QOL) in clinically stable patients with COVID-19. METHODS: This was an online survey conducted in COVID-19 patients across five designated isolation hospitals for COVID-19 in Hubei province, China. Depression and QOL were assessed with standardized instruments. RESULTS: A total of 770 participants were included. The prevalence of depression was 43.1% (95%CI: 39.6%-46.6%). Binary logistic regression analysis found that having a family member infected with COVID-19 (OR=1.51, P = 0.01), suffering from severe COVID-19 infection (OR=1.67, P = 0.03), male gender (OR=0.53, P<0.01), and frequent social media use to obtain COVID-19 related information (OR=0.65, P<0.01) were independently associated with depression. Patients with depression had lower QOL than those without. CONCLUSION: Depression is highly prevalent in clinically stable patients with COVID-19. Regular screening and appropriate treatment of depression are urgently warranted for this population.

Statistical and network analysis of 1212 COVID-19 patients in Henan, China.

BACKGROUND: COVID-19 is spreading quickly all over the world. Publicly released data for 1212 COVID-19 patients in Henan of China were analyzed in this paper. METHODS: Various statistical and network analysis methods were employed. RESULTS: We found that COVID-19 patients show gender (55% vs 45%) and age (81% aged between 21 and 60) preferences; possible causes were explored. The estimated average, mode and median incubation periods are 7.4, 4 and 7 days. Incubation periods of 92% of patients were no more than 14 days. The epidemic in Henan has undergone three stages and has shown high correlations with the numbers of patients recently returned from Wuhan. Network analysis revealed that 208 cases were clustering infected, and various People's Hospitals are the main force in treating COVID-19. CONCLUSIONS: The incubation period was statistically estimated, and the proposed state transition diagram can explore the epidemic stages of emerging infectious disease. We suggest that although the quarantine measures are gradually working, strong measures still might be needed for a period of time, since ∼7.45% of patients may have very long incubation periods. Migrant workers or college students are at high risk. State transition diagrams can help us to recognize the time-phased nature of the epidemic. Our investigations have implications for the prevention and control of COVID-19 in other regions of the world.